幽门螺杆菌对甲硝唑耐药性及多重相关基因DNA和氨基酸序列分析

Susceptibilities of Helicobacter pylori Strains to Metronidazole and the Analysis of Multiple Related Genes Sequences of DNA and Amino Acids

目的 了解幽门螺杆菌(Helicobacter pylori, Hp) 对甲硝唑的耐药状况, 研究相关的多重基因突变, 为深入揭示Hp耐药的分子机制提供相关资料。方法 分离培养54株Hp, 以琼脂稀释法检测Hp的甲硝唑最低抑菌浓度(deterimination of minimal inhibitory concentration, MIC), 用PCR方法扩增甲硝唑耐药相关基因(rdxA、frxA、fdxB), 经分子克隆后DNA测序, 分析敏感株和耐药株耐药基因的序列及其与耐药性的关系。结果 武汉地区人群Hp对甲硝唑的耐药率为67%; 甲硝唑耐药基因序列分析表明, 耐药临床株的rdxA和(或) frxA基因存在插入、缺失和点突变。结论 Hp对甲硝唑耐药不仅与rdxA有关, 同时存在frxA、fdxB基因突变。frxA、fdxB基因的突变可能与rdxA在Hp耐甲硝唑中有协同作用。

Objective To investigate the resistance of Helicobacter pylori (H. pylori) to metronidazole and analyze related genes rdxA,frxA and fdxB for providing data used for understanding molecular mechanisms of the drug resistance of the pathogens.Methods H. pylori were isolated from gastric mucosa biopsies of patients with peptic ulcer and gastritis. Agar-diffusion method was used for testing susceptibility of H. pylori to metronidazole. Total DNAs of the pathogens were then isolated and purified for amplification of genes rdxA,frxA and fdxB by PCR. Target DNAs were cloned and sequenced and then analyzed for the relationship between gene mutations and the metronidazole resistance.Results The prevalence of antibiotic resistance to metronidazole in H. pylori clinical isolates were 67 % in Hubei area. DNA sequencing showed that insertion,deletion and point mutations existed in the genes rdxA,frxA,fdxB.Conclusion Mutations of rdxA and frxA simultaneously occurred in metronidazole resistant strains,implicating that the gene frxA is likely cooperative to effect of rdxA in resistance of H. pylori to metronidazole.